IDH1 mutation produces R-2-hydroxyglutarate (R-2HG) and induces mir-182-5p expression to regulate cell cycle and tumor formation in glioma.

BACKGROUND: Mutations in isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2), are present in most gliomas. IDH1 mutation is an important prognostic marker in glioma. However, its regulatory mechanism in glioma remains incompletely understood. RESULTS: miR-182-5p expression was increased within IDH1-mutant glioma specimens according to TCGA, CGGA, and online dataset GSE119740, as well as collected clinical samples. (R)-2-hydroxyglutarate ((R)-2HG) treatment up-regulated the expression of miR-182-5p, enhanced glioma cell proliferation, and suppressed apoptosis; miR-182-5p inhibition partially eliminated the oncogenic effects of R-2HG upon glioma cells. By direct binding to Cyclin Dependent Kinase Inhibitor 2 C (CDKN2C) 3'UTR, miR-182-5p inhibited CDKN2C expression. Regarding cellular functions, CDKN2C knockdown promoted R-2HG-treated glioma cell viability, suppressed apoptosis, and relieved cell cycle arrest. Furthermore, CDKN2C knockdown partially attenuated the effects of miR-182-5p inhibition on cell phenotypes. Moreover, CDKN2C knockdown exerted opposite effects on cell cycle check point and apoptosis markers to those of miR-182-5p inhibition; also, CDKN2C knockdown partially attenuated the functions of miR-182-5p inhibition in cell cycle check point and apoptosis markers. The engineered CS-NPs (antagomir-182-5p) effectively encapsulated and delivered antagomir-182-5p, enhancing anti-tumor efficacy in vivo, indicating the therapeutic potential of CS-NPs(antagomir-182-5p) in targeting the miR-182-5p/CDKN2C axis against R-2HG-driven oncogenesis in mice models. CONCLUSIONS: These insights highlight the potential of CS-NPs(antagomir-182-5p) to target the miR-182-5p/CDKN2C axis, offering a promising therapeutic avenue against R-2HG's oncogenic influence to glioma.

Transcriptomic analysis identifies the neuropeptide cortistatin (CORT) as an inhibitor of temozolomide (TMZ) resistance by suppressing the NF-κB-MGMT signaling axis in human glioma.

Glioma is a common tumor originating in the brain that has a high mortality rate. Temozolomide (TMZ) is the first-line treatment for high-grade gliomas. However, a large proportion of gliomas are resistant to TMZ, posing a great challenge to their treatment. In the study, the specific functions and mechanism(s) by which cortistatin (CORT) regulates TMZ resistance and glioma progression were evaluated. The decreased expression of CORT was detected in glioma tissues, and highly expressed CORT was associated with a better survival rate in patients with glioma. CORT overexpression notably decreased the capacity of glioma cells to proliferate and migrate in vitro and to form tumors in vivo. CORT overexpression also markedly suppressed the viability and enhanced the apoptosis of TMZ-resistant U251 cells by regulating MGMT, p21, and Puma expression. Importantly, CORT overexpression reduced the resistance of gliomas to TMZ in vivo. CORT expression was negatively correlated with MGMT expression in both glioma tissues and cells, and it was found that CORT inhibited NF-κB pathway activation in glioma cells, thereby inhibiting MGMT expression. In conclusion, CORT regulates glioma cell growth, migration, apoptosis, and TMZ resistance by weakening the activity of NF-κB/p65 and thereby regulating MGMT expression. The CORT/NF-κB/MGMT axis might be regarded as a molecular mechanism contributing to the resistance of glioma to TMZ. Our data also suggest that CORT regulates the viability and metastatic potential of glioma cells, independent of its effects on TMZ resistance, providing evidence of novel therapeutic targets for glioma that should be evaluated in further studies.

Decrease of Cellular Communication Network Factor 1 (CCN1) Attenuates PTZ-Kindled Epilepsy in Mice.

To investigate the molecular mechanism of communication network factor 1 (CCN1) regulating pentylenetetrazol (PTZ)-induced epileptogenesis, deepen the understanding of epilepsy seizure pathogenesis, and provide new drug action targets for its clinical prevention and treatment. Differentially expressed genes (DEGs) on microarrays GSE47516 and GSE88992 were analyzed online using GEO2R. Pathway enrichment and protein-protein interaction network (PPI) analysis of DEGs were carried out using Metascape. Brain tissue samples of severe traumatic brain injury patients (named Healthy group) and refractory epilepsy patients (named Epilepsy group) were obtained and analyzed by qRT-PCR and immunohistochemistry (IHC) staining. A PTZ-induced epilepsy mouse model was established and verified. Morphological changes of neurons in mouse brain tissue were detected using hematoxylin and eosin (HE) staining. qRT-PCR was conducted to detect the mRNA expressions of apoptosis-associated proteins Bax, Caspase-3 and bcl2. TUNEL staining was performed to detect brain neuron apoptosis. The levels of myocardial enzymology, GSH, MDA and ROS in blood of mouse were detected by biochemical assay. CCN1 expression was increased in epilepsy brain tissue samples. CCN1 decreasing effectively prolongs seizure incubation period and decreases seizure duration. Silencing of CCN1 also reduces neuronal damage and apoptosis, decreases mRNA and protein expression of proapoptotic proteins Bax and Caspase-3, increases mRNA expression of antiapoptotic protein Bcl2. Moreover, decrease of CCN1 decreases myocardial enzymatic indexes CK and CK-MB levels, reduces myocardial tissue hemorrhage, and relieves oxidative stress response in hippocampal and myocardial tissue. CCN1 expression is increased in epileptic samples. CCN1 decreasing protects brain tissue by attenuating oxidative stress and inhibiting neuronal apoptosis triggered by PTZ injection, which probably by regulating Nrf2/HO-1 pathway.

Activation of Epac in the BLA disrupts reconsolidation and attenuates heroin-seeking behaviour.

The susceptibility to drug cravings evoked by stimuli poses a formidable hurdle in the treatment of addiction and the prevention of relapse. Pharmacological interventions targeting drug-associated memories hold promise for curbing relapse by impeding the process of memory reconsolidation, predominantly governed by cAMP signalling. Exchange Protein Activated by cAMP (Epac) serves as a distinctive mediator of cAMP signalling, which has been implicated in reinforcing the effects of cocaine and facilitating the acquisition. Nonetheless, the role of Epac in heroin-related memory and the subsequent seeking behaviour remains enigmatic. In this study, we explored the impact of Epac activation on the reconsolidation process of drug-related memories associated with heroin self-administration. Over the course of 10 consecutive days, rats underwent training, wherein they acquired the behaviour of nose poking to obtain heroin accompanied by a tone + light cue. This nose-poking behaviour was subsequently extinguished when heroin infusion and cue presentation were discontinued. Subsequently, we administered 8-pCPT-cAMP (8-CPT), an Epac-specific activator, into the basolateral amygdala at various time points, either in the presence or absence of a conditioned stimulus. Our findings demonstrate that administering 8-CPT immediately after memory retrieval effectively reduces cue- and heroin-induced reinstatement, with the observed effects persisting significantly for a minimum of 28 days. However, infusion of 8-CPT for a duration of 6 h following the memory retrieval trial, or without it altogether, had no discernible impact. Thus, these findings strongly suggest that Epac activation can disrupt the reconsolidation of heroin-associated memory, thereby diminishing the reinstatement of heroin-seeking behaviour.

The Cdk5 inhibitor ß-butyrolactone impairs reconsolidation of heroin-associated memory in the rat basolateral amygdala.

The persistence of maladaptive heroin-associated memory, which is triggered by drug-related stimuli that remind the individual of the drug's pleasurable and rewarding effects, can impede abstinence efforts. Cyclin-dependent kinase 5 (Cdk5), a neuronal serine/threonine protein kinase that plays a role in multiple neuronal functions, has been demonstrated to be involved in drug addiction and learning and memory. Here, we aimed to investigate the role of cdk5 activity in the basolateral amygdala (BLA) in relapse to heroin seeking, using a self-administration rat model. Male rats underwent 10 days of heroin self-administration training, during which an active nose poke resulted in an intravenous infusion of heroin that was accompanied by a cue. The rats then underwent nose poke extinction for 10 days, followed by subsequent tests of heroin-seeking behaviour. We found that intra-BLA infusion of ß-butyrolactone (100 ng/side), a Cdk5 inhibitor, administered 5 min after reactivation, led to a subsequent decrease in heroin-seeking behaviour. Further experiments demonstrated that the effects of ß-butyrolactone are dependent on reactivated memories, temporal-specific and long-lasting on relapse of heroin-associated memory. Results provide suggestive evidence that the activity of Cdk5 in BLA is critical for heroin-associated memory and that the specific inhibitor, ß-butyrolactone, may hold potential as a substance for the treatment of heroin abuse.

The miR-27a-3p/FTO axis modifies hypoxia-induced malignant behaviors of glioma cells.

Glioblastoma multiforme (GBM) is one of the most malignant types of central nervous system (CNS) tumors. N6-methyladenine (m6A) RNA modification is a main type of RNA modification in eukaryotic cells. In this study, we find that the m6A RNA methylation eraser FTO is dramatically downregulated in glioma samples and cell lines, particularly in intermediate and core regions and hypoxia-challenged glioma cells. In vitro, FTO overexpression inhibits the hypoxia-induced capacities of glioma cells to proliferate, migrate and invade, and decreases the percentage of cells with m6A RNA methylation. In vivo, FTO overexpression inhibits tumor growth in the xenograft model and decreases the protein levels of migration markers, including Vimentin and Twist. miR-27a-3p is upregulated within glioma intermediate and core regions and hypoxia-challenged glioma cells. miR-27a-3p inhibits the expression of FTO via direct binding to FTO. miR-27a-3p overexpression promotes hypoxia-challenged glioma cell aggressiveness, whereas FTO overexpression partially diminishes the oncogenic effects of miR-27a-3p overexpression. FTO overexpression promotes the nuclear translocation of FOXO3a and upregulates the expression levels of the FOXO3a downstream targets BIM, BNIP3, BCL-6, and PUMA, possibly by interacting with FOXO3a. Conclusively, FTO serves as a tumor suppressor in glioma by suppressing hypoxia-induced malignant behaviors of glioma cells, possibly by promoting the nuclear translocation of FOXO3a and upregulating FOXO3a downstream targets. miR-27a-3p is a major contributor to FTO downregulation in glioma under hypoxia.

Identification of tryptophan metabolic gene-related subtypes, development of prognostic models, and characterization of tumor microenvironment infiltration in gliomas.

Background: Epigenetic regulation and immunotherapy of tumor microenvironment (TME) is a hot topic in recent years. However, the potential value of tryptophan metabolism genes in regulating TME and immunotherapy is still unclear. Materials and methods: A comprehensive study of glioma patients was carried out based on 40 tryptophan metabolic genes. Subsequently, these prognostic tryptophan metabolic genes are systematically associated with immunological characteristics and immunotherapy. A risk score model was constructed and verified in the Cancer Genome Atlas (TCGA) and the Chinese Glioma Genome Atlas (CGGA) cohorts to provide guidance for prognosis prediction and immunotherapy of glioma patients. Results: We described the changes of tryptophan metabolism genes in 966 glioma samples from genetic and transcriptional fields and evaluated their expression patterns from two independent data sets. We identified two different molecular subtypes and found that two subtypes were associated with clinicopathological features, prognosis, TME cell infiltration, and immune checkpoint blockers (ICBs). Then, four genes (IL4I1, CYP1A1, OGDHL, and ASMT) were screened out by univariate and multivariate cox regression analysis of tryptophan metabolism genes, and a risk score model for predicting the overall survival (OS) of glioma patients was constructed. And its predictive ability is verified using the CGGA database. At the same time, we verified the expression of IL4I1, CYP1A1, OGDHL, and ASMT four genes in glioma specimens and cell lines in GES4260 and GES15824. Therefore, we constructed a nomogram to improve the clinical applicability of the risk assessment model. The high risk score group, characterized by increased TMB and immune cell infiltration, was also sensitive to temozolomide immunotherapy. Our comprehensive analysis of tryptophan metabolic genes in gliomas shows that they play a potential role in tumor immune stromal microenvironment, clinicopathological features, and prognosis. Conclusion: Tryptophan metabolism genes play an indispensable role in the complexity, diversity, and prognosis of TME. This risk score model based on tryptophan metabolism gene is a new predictor of clinical prognosis and immunotherapy response of glioma, and guides a more appropriate immunotherapy strategy for glioma patients.

Mechanisms for hypoxia-induced long non-coding small nucleolar RNA host gene 14 in promoting temozolomide resistance of glioma. / ä½Žæ°§è¯±å¯¼é•¿é“¾éžç¼–ç æ ¸å† å°RNAå®¿ä¸»åŸºå› 14促进胶质瘤替莫唑胺耐药的机制.

OBJECTIVES: This study aims to investigate the role of hypoxia-induced long non-coding small nucleolar RNA host gene 14 (lncRNA SNHG14) in glioma temozolomide (TMZ) resistance and underlying mechanisms. METHODS: According to different treatments, the experiment was divided into a normoxia group and a hypoxia group, a control group and a TMZ group. The lncRNA SNHG14 and O6-methylguanine DNA methyltransferase (MGMT) levels in glioma SNB19 and U251 cell line were detected by real-time PCR and Western blotting, respectively, and the association of lncRNA SNHG14 level with hypoxia and TMZ treatment was analyzed. siRNA was used to knockdown the lncRNA SNHG14 expression in glioma cells, and the transfected glioma cells were divided into a negative control group (si-NC group) and a si-SNHG14 group. The interference efficiency was examined by real-time PCR, the key factor MGMT of lncRNA SNHG14 sensitivity regulation was detected by Western blotting, and the cell apoptosis was detected by cytometry. In addition, MTT method was used to detect the cell viability of gliomas in the different groups under the different TMZ concentrations, and the effect of lncRNA SNHG14 on TMZ sensitivity of gliomas was analyzed. Online tools were used to predict miRNAs that could specifically bind to lncRNAs SNHG14 and MGMT. A si-NC group, a si-SNHG14 group, a normoxia group and a hypoxia group were set up, and the changes of miR-143 abundance in different environments were observed by real-time PCR. miR-143 mimics and inhibitor were used to change the level of miR-143 in glioma cells. A NC inhibitor group, a miR-143 inhibitor group, a NC mimics group and a miR-143 mimics group were set up, the interference efficiency was detected by real-time PCR, the expression level of MGMT was detected by Western blotting, and the effect of miR-143 on the level of MGMT were analyzed. The NC inhibitor group, the miR-143 inhibitor group, the NC mimics group and the miR-143 mimics group were treated with different interventions, and the dual luciferase reporter assay was used to observe the changes of lncRNA SNHG14 and MGMT luciferase activities, and to verify the relationship among lncRNA SNHG14, miR-143 and MGMT. Finally, a NC group and a lncRNA SNHG14 overexpression group were set up, and the changes in the abundance of miR-143 and MGMT in each group were detected by RNA-binding protein immunoprecipitation experiments, and the competitive binding relationship among lncRNA SNHG14, miR-143 and MGMT was analyzed. RESULTS: Compared with the normoxia group, the hypoxia group could promote the expression of lncRNA SNHG14 in glioma cells. Compared with the control group, the expression of lncRNA SNHG14 could be significantly inhibited in the TMZ group (P<0.05). Compared with the si-NC group, the expression of lncRNA SNHG14 in the si-SNHG14 group could be effectively inhibited, and the expression level of MGMT was significantly decreased, and the apoptosis rate was significantly increased (all P<0.05). With the increase of TMZ concentrations, the glioma cell viability in the si-SNHG14 group was significantly lower than that in the si-NC group, and the cell viability in the hypoxia group was significantly higher than that in the normoxia group (both P<0.05). Online tool prediction found that miR-143 had binding sites with lncRNA SNHG14 and MGMT. The abundance of miR-143 in the hypoxia group was significantly lower than that in the normoxic group, and the abundance of miR-143 in the si-SNHG14 group was significantly higher than that in the si-NC group (both P<0.05). The miR-143 mimics group or the miR-143 inhibitor group could significantly over-express or under-express miR-143 (both P<0.05). But there was no significant difference between the NC mimics group (or the NC inhibitor group) and the control group (both P>0.05). The level of MGMT protein could significantly up-regulate in the miR-143 inhibitor group, and on the contrary which could significantly down-regulate in the miR-143 mimics group (both P<0.01). The dual luciferase reporter assay showed that there was no significant difference between the NC mimics group (or the NC inhibitor group) and the control group (both P>0.05). The wild-type SNHG14 and MGMT luciferase activities were significantly down-regulated in the miR-143 mimics group, which were significantly up-regulated in the miR-143 inhibitor group (P<0.01 and P<0.05, respectively), but there was no significant change in the luciferase activities of mutant SNHG14 and MGMT (both P>0.05). The results of the RNA-binding protein immunoprecipitation experiment showed that: compared with the NC group, more lncRNA SNHG14 was bound to the precipitated argonaute 2 protein in the cells in the lncRNA SNHG14 overexpression group, but the abundance of MGMT mRNA was decreased significantly, and there were significant differences (both P<0.01). There was a targeting regulatory relationship among lncRNA SNHG14, miR-143 and MGMT. CONCLUSIONS: The up-regulated lncRNA SNHG14 can target miR-143, relieve the inhibition of miR-143 on MGMT, and promote the TMZ resistance in the hypoxia-induced glioma cells.

Long Noncoding RNA OR7E156P/miR-143/HIF1A Axis Modulates the Malignant Behaviors of Glioma Cell and Tumor Growth in Mice.

Gliomas are characterized by high incidence, recurrence and mortality all of which are significant challenges to efficacious clinical treatment. The hypoxic microenvironment in the inner core and intermediate layer of the tumor mass of gliomas is a critical contributor to glioma pathogenesis. In this study, we identified an upregulated lncRNA, OR7E156P, in glioma was identified. The silencing of OR7E156P inhibited cell invasion and DNA synthesis in vitro and tumor growth in vivo. OR7E156P was intricately linked to the HIF1A pathway. Hypoxia could induce OR7E156P expression, whereas OR7E156P silencing decreased HIF1A protein levels under hypoxic conditions. Hypoxia promoted glioma cell invasion and DNA synthesis, and HUVEC tube formation, whereas OR7E156P silencing partially reversed the cellular effects of hypoxia. HIF1A overexpression promoted, whereas OR7E156P silencing inhibited tumor growth; the inhibitory effects of OR7E156P silencing on tumor growth were partially reversed by HIF1A overexpression. miR-143 directly targeted OR7E156P and HIF1A, respectively. miR-143 inhibition increased HIF1A protein levels, promoted glioma cell invasion and DNA synthesis. Moreover, they enhanced HUVEC tube formation, whereas OR7E156P silencing partially reversed the cellular effects of miR-143 inhibition. HIF1A targeted the promoter region of miR-143 and inhibited miR-143 expression. Altogether a regulatory axis consisting of OR7E156P, miR-143, and HIF1A, was identified which is deregulated in glioma, and the process of the OR7E156P/miR-143/HIF1A axis modulating glioma cell invasion through ZEB1 and HUVEC tube formation through VEGF was demonstrated.

CircRNA NALCN acts as an miR-493-3p sponge to regulate PTEN expression and inhibit glioma progression.

BACKGROUND: An increasing number of studies have shown that circular RNAs (circRNAs) play important roles in the regulation of tumor progression. Therefore, we explored the expression characteristics, function, and related mechanism of the newly identified circNALCN in glioma. METHODS: RNA sequencing was used to analyze the expression profiles of circRNAs in brain tissue from five glioma cases and four normal controls. Quantitative real-time polymerase chain reaction was implemented to examine the levels of circNALCN, miR-493-3p, and phosphatase and tensin homolog (PTEN). Cell counting kit 8 assays were performed to analyze cell proliferation, and cell migration was assessed by the wound healing test and Transwell assay. Dual-luciferase reporter, fluorescence in situ hybridization, and RNA pulldown assays were performed to confirm the role of circNALCN as an miR-493-3p sponge, weakening the inhibitory effect of miR-493-3p on target PTEN expression. RESULTS: The downregulated expression of circNALCN was observed in both glioma tissues and cell lines. CircNALCN expression was negatively correlated with World Health Organization grade and overall survival in patients with glioma. Functionally, the overexpression of circNALCN significantly inhibited the proliferation and migration of glioma cells, whereas miR-493-3p mimics counteracted these effects. The mechanistic analysis demonstrated that circNALCN acted as a competing endogenous RNA for miR-493-3p to relieve the repressive effects of miR-493-3p on its target, PTEN, suppressing glioma tumorigenesis. CONCLUSIONS: CircNALCN inhibits the progression of glioma through the miR-493-3p/PTEN axis, providing a developable biomarker and therapeutic target for glioma patients.

A HIF1A/miR-485-5p/SRPK1 axis modulates the aggressiveness of glioma cells upon hypoxia.

The high aggressiveness of gliomas remains a huge challenge to clinical therapies, and the hypoxic microenvironment in the core region is a critical contributor to glioma aggressiveness. In this study, it was found that miR-485-5p was low expressed within glioma tissue samples and cells. GO enrichment annotation indicated that the predicted downstream targets miR-485-5p were enriched in hypoxia response and decreased oxygen level. In glioma cells, miR-485-5p overexpression suppressed cell viability, migratory ability, and invasive ability under both normoxic and hypoxic conditions. Through direct binding, miR-485-5p suppressed SRPK1 expression. Under hypoxia, SRPK1 overexpression enhanced hypoxia-induced glioma cell aggressiveness and significantly reversed the effects of miR-485-5p overexpression. Moreover, HIF1A could target the miR-485-5p promoter region to inhibit the transcription. HIF1A, miR-485-5p, and SRPK1 form a regulatory axis, which modulates glioma cell aggressiveness under hypoxia. In conclusion, we identify a HIF1A/miR-485-5p/SRPK1 axis that modulates the aggressiveness of glioma cells under hypoxia. The axis could potentially provide new research avenues in the treatment of gliomas considering the hypoxic environment in its core.

Graphite Felt Modified by Atomic Layer Deposition with TiO2 Nanocoating Exhibits Super-Hydrophilicity, Low Charge-Transform Resistance, and High Electrochemical Activity.

Graphite felt (GF) is a multi-functional material and is widely used as electrodes of electrochemical devices for energy and environmental applications. However, due to the inherent hydrophobicity of graphite felt, it must be hydrophilically pretreated to obtain good electrochemical activity. Metal oxides coating is one of the feasible methods to modify the surface of GF, and in order to ensure that the metal oxides have a better conductivity for obtaining higher electrochemical activity, a subsequent H2 heat-treatment process is usually adopted. In this study, atomic layer deposition (ALD) is used to deposit TiO2 nanocoating on graphite felt (GF) for surface modification without any H2 thermal post-treatment. The results show that the ALD-TiO2-modified GF (ALD-TiO2/GF) owns excellent hydrophilicity. Moreover, the ALD-TiO2/GF exhibits excellent electrochemical properties of low equivalent series resistance (Rs), low charge-transfer resistance (Rct), and high electrochemical activity. It demonstrates that ALD is an applicable technique for modifying the GF surface. In addition, it can be reasonably imagined that not only TiO2 film can effectively modify the GF surface, but also other metal oxides grown by ALD with nanoscale-thickness can also obtain the same benefits. We anticipate this work to be a starting point for modifying GF surface by using ALD with metal oxides nanocoating.

ANXA2P2/miR-9/LDHA axis regulates Warburg effect and affects glioblastoma proliferation and apoptosis.

BACKGROUND: Aerobic glycolysis is a unique tumor cell phenotype considered as one of the hallmarks of cancer. Aerobic glycolysis can accelerate tumor development by increasing glucose uptake and lactate production. In the present study, lactate dehydrogenase A (LDHA) is significantly increased within glioma tissue samples and cells, further confirming the oncogenic role of LDHA within glioma. METHODS: Hematoxylin and eosin (H&E) and immunohistochemical (IHC) staining were applied for histopathological examination. The protein levels of LDHA, transporter isoform 1 (GLUT1), hexokinase 2 (HK2), phosphofructokinase (PFK) in target cells were detected by Immunoblotting. The predicted miR-9 binding to lncRNA Annexin A2 Pseudogene 2 (ANXA2P2) or the 3' untranslated region (UTR) of LDHA was verified using Luciferase reporter assay. Cell viability or apoptosis were examined by MTT assay or Flow cytometry. Intracellular glucose and Lactate levels were measured using glucose assay kit and lactate colorimetric assay kit. RESULTS: The expression of ANXA2P2 showed to be dramatically upregulated within glioma tissue samples and cells. Knocking down ANXA2P2 within glioma cells significantly inhibited cell proliferation and aerobic glycolysis, as manifested as decreased lactate and increased glucose in culture medium, and downregulated protein levels of glycolysis markers, GLUT1, HK2, PFK, as well as LDHA. miR-9 was predicted to target both lncRNA ANXA2P2 and LDHA. The overexpression of miR-9 suppressed the cell proliferation and aerobic glycolysis of glioma cells. Notably, miR-9 could directly bind to LDHA 3'UTR to inhibit LDHA expression and decrease the protein levels of LDHA. ANXA2P2 competitively targeted miR-9, therefore counteracting miR-9-mediated repression on LDHA. Within tissues, miR-9 exhibited a negative correlation with ANXA2P2 and LDHA, respectively, whereas ANXA2P2 and LDHA exhibited a positive correlation with each other. CONCLUSIONS: In conclusion, ANXA2P2/miR-9/LDHA axis modulates the aerobic glycolysis progression in glioma cells, therefore affecting glioma cell proliferation.

Etomidate Effects on Desensitization and Deactivation of α4ß3δ GABAA Receptors Inducibly Expressed in HEK293 TetR Cells.

Central α4ßδ receptors are the most abundant isoform of δ subunit-containing extrasynaptic GABAA receptors that mediate tonic inhibition. Although the amplitude of GABA-activated currents through α4ßδ receptors is modulated by multiple general anesthetics, the effects of general anesthetics on desensitization and deactivation of α4ßδ receptors remain unknown. In the current study, we investigated the effect of etomidate, a potent general anesthetic, on the kinetics and the pseudo steady-state current amplitude of α4ß3δ receptors inducibly expressed in human embryonic kidney 293 TetR cells. Etomidate directly activates α4ß3δ receptors in a concentration-dependent manner. Etomidate at a clinically relevant concentration (3.2 µM) enhances maximal response without altering the EC50 of GABA concentration response. Etomidate also increases the extent of desensitization and prolongs the deactivation of α4ß3δ receptors in the presence of maximally activating concentrations of GABA (1 mM). To mimic the modulatory effect of etomidate on tonic currents, long pulses (30-60 seconds) of a low GABA concentration (1 µM) were applied to activate α4ß3δ receptors in the absence and presence of etomidate. Although etomidate increases the desensitization of α4ß3δ receptors, the pseudo steady-state current amplitude at 1 µM GABA is augmented by etomidate. Our data demonstrate that etomidate enhances the pseudo steady-state current of α4ß3δ receptors evoked by a GABA concentration comparable to an ambient GABA level, suggesting that α4ß3δ receptors may mediate etomidate's anesthetic effect in the brain.

MicroRNA-509-3p inhibits cell proliferation and invasion via downregulation of X-linked inhibitor of apoptosis in glioma.

Malignant glioma is an aggressive type of cancer. Increasing evidence has suggested that microRNAs (miRs) regulate gene expression post-transcriptionally to affect cancer development and progression. Aberrant expression of miR-509-3p has been reported in cancer studies. However, the expression and mechanism of its function in glioma remains unclear. The present study demonstrated that miR-509-3p was downregulated in glioma tissue samples relative to non-tumor tissues, and that low miR-509-3p expression was associated with a reduced overall survival time. Functional studies revealed that the overexpression of miR-509-3p inhibited cell proliferation, induced apoptosis and suppressed cell migration and invasion via negatively regulating the expression of X-linked inhibitor of apoptosis. The data therefore suggested that miR-509-3p serves an important role in the development and progression of glioma, implicating its possible application in clinical practice as a biomarker and a potential novel therapeutic target.

NGF/FAK signal pathway is implicated in angiogenesis after acute cerebral ischemia in rats.

Neurogenesis in the cerebral infarction after an ischemic event is important to the rehabilitation of patients. However, the mechanism of angiogenesis around cerebral ischemia is not clear. Our study designed to test whether the nerve growth factor (NGF)-P-focal adhesion kinase (FAK) signaling pathway for associations with angiogenesis plays a key role in post-acute cerebral ischemia of rats. Firstly, we implanted the Matrigel, a carrier of basement membrane matrix, into the abdominal skin of rats to identify the relevant components of the NGF-P-FAK signaling pathway related to angiogenesis. Secondly, we used a model established by ligation of the middle cerebral artery (MCA) to observe the effect of the same signal pathway on angiogenesis in the subventricular and subgranular zones of the dentate gyrus(SVG and SGZ). The results showed that the tissue scores was significantly increased by NGF. However, the tissue scores was signifcaintly decreased by FAK inhibitor TAE226. Furthermore, CD31 and α-SMA were significantly increased by NGF and were decreased by anti-NGF and TAE226 in Matrigel. The P-FAK protein expression in Matrigel was markedly increased by NGF and decreased by TAE226. In the SVZ and SVG of cerebral ischemia, the numbers of BrdU-positive cells were significantly increased by NGF and decreased by TAE226, respectively. Our findings suggest that the therapy targeting the NGF-P-FAK signaling pathway may be an option for patients suffering from cerebral ischemia.

Pathophysiological changes in the cerebellum and brain stem in a rabbit model after superior petrosal vein sacrifice.

In certain surgical procedures, sacrificing the superior petrosal vein (SPV) is required. Previous studies have reported transient cerebellar edema, venous infarction, or hemorrhage that might occur after sectioning of the SPV. The present study investigated the pathophysiological changes in cerebellum and brain stem after SPV sacrifice. Rabbits were divided into the operation group where the SPV was sacrificed and the control group where the SPV remained intact. Each group was further subdivided into 4, 8, 12, 24, 48, and 72 h groups which represented the time period from sacrificing of the SPV to killing of the rabbits. The water content (WC), Na+ content, K+ content, and pathophysiological changes in cerebellum and brain stem tissue were measured. In comparison with the control, the WC and Na+ content of cerebellar tissue were increased in the 4, 8, 12, and 24 h operation subgroups (P<0.05), but only increased in the 4-h subgroup of the brain stem tissue (P<0.05). The K+ content of the cerebellar tissue decreased in the 4, 8, 12, and 24 h operation subgroups (P<0.05) but only decreased in the 4-h subgroup of brain stem tissue (P<0.05). Nissl staining and TEM demonstrated that cerebellar edema occurred in the 4, 8, 12, and 24 h operation subgroups but not in the 48- and 72-h subgroups. Brain stem edema occurred in the 4-h operation subgroup. In summary, cerebellum and brain stem edema can be observed at different time points after sacrificing of the SPV in the rabbit model.

BAF53a is a potential prognostic biomarker and promotes invasion and epithelial-mesenchymal transition of glioma cells.

Increasing evidence indicates that BAF53a is crucial for embryonic development and maintenance of stemness, and may be associated with epithelial-mesenchymal transition (EMT), which suggests its involvement in cancer progression. However, the role of BAF53a in glioma remains unknown. In the present study, BAF53a was found to be highly expressed in glioma tissues and was associated with poor overall survival (OS) and progression-free survival (PFS) in glioma patients. A multivariate Cox regression analysis revealed that BAF53a might be an independent prognostic factor for OS and PFS in glioma patients. Further functional analysis indicated that BAF53a overexpression could promote proliferation and increase the motility and invasion of U87 glioma cells, whereas BAF53a knockdown had the opposite effect. In addition, BAF53a expression was associated with the levels of E­cadherin and vimentin expression in glioma tissues. This was further confirmed in U87 cells expressing different levels of BAF53a; BAF53a overexpression was concomitant with decreased E­cadherin and increased vimentin expression, whereas BAF53a knockdown showed the opposite pattern of expression. Taken together, these results suggest that BAF53a may be a novel prognostic factor for glioma patients, and that BAF53 may facilitate glioma progression by promoting proliferation, invasion, and associate with EMT. Therefore, BAF53a could be a potential promising biomarker and a target for the treatment of glioma.

LncRNA-XIST interacts with miR-29c to modulate the chemoresistance of glioma cell to TMZ through DNA mismatch repair pathway.

Temozolomide (TMZ) is the most commonly used alkylating agent in glioma chemotherapy. However, growing resistance to TMZ remains a major challenge for clinicians. Recent evidence emphasizes the key regulatory roles of non-coding RNAs (lncRNAs and miRNAs) in tumor biology, including the chemoresistance of cancers. However, little is known about the role and regulation mechanisms of lncRNA cancer X-inactive specific transcripts (XIST) in glioma tumorigenesis and chemotherapy resistance. In the present study, higher XIST expression was observed in glioma tissues and cell lines, which was related to poorer clinicopathologic features and shorter survival time. XIST knockdown alone was sufficient to inhibit glioma cell proliferation and to amplify TMZ-induced cell proliferation inhibition. Moreover, XIST knockdown can sensitize TMZ-resistant glioma cells to TMZ. XIST can inhibit miR-29c expression by directly targetting TMZ-resistant glioma cells. DNA repair protein O6-methylguanine-DNA methytransferase (MGMT) plays a key role in TMZ resistance; transcription factor specificity protein 1 (SP1), a regulator of DNA mismatch repair (MMR) key protein MSH6, has been reported to be up-regulated in TMZ-resistant glioma cell lines. In the present study, we show that XIST/miR-29c coregulates SP1 and MGMT expression in TMZ-resistant glioma cell lines. Our data suggest that XIST can amplify the chemoresistance of glioma cell lines to TMZ through directly targetting miR-29c via SP1 and MGMT. XIST/miR-29c may be a potential therapeutic target for glioma treatment.

Effect of TNF-α Inhibition on Bone Marrow-Derived Mesenchymal Stem Cells in Neurological Function Recovery after Spinal Cord Injury via the Wnt Signaling Pathway in a Rat Model.

AIM: The present study aimed to examine the effect of tumor necrosis factor-α (TNF-α) inhibition on bone marrow-derived mesenchymal stem cells (BMSCs) in neurological function recovery after spinal cord injury (SCI) via the Wnt signaling pathway in a rat model. METHODS: The rat model of SCI was established using Allen's method. Seventy-two adult male Sprague Dawley (SD) rats were randomly assigned into 4 groups (18 rats in each group): the sham control group, saline control group, BMSCs group (injection with BMSCs at the injured site) and BMSCs + TNF-α group (injection with BMSCs under TNF-α treatment at the injured site). Immunochemistry was performed to characterize the culture media after TNF-α-induced differentiation. qRT-PCR and Western blotting analyses were performed to detect the mRNA and protein expression of ß-catenin, Wnt3a, GSK-3ß and Axin. The Basso Beattie Bresnahan (BBB) locomotor score, neurological deficit score (NDS), and balance beam test (BBT) score were used to assess neurological functional recovery of SCI rats. RESULTS: In the BMSC group, numerous spherical cell clusters grew in suspension, and the cells were nestin-, NF200- and GFAP-positive. Compared with the sham control and BMSC groups, the ß-catenin and Wnt3a mRNA and protein expression was increased, but the GSK-3ß and Axin mRNA and protein expression was decreased in the BMSCs + TNF-α group. The SCI rats in the BMSCs + TNF-α group exhibited lower BBB scores, and higher NDSs and BBT scores compared to the BMSCs group. CONCLUSION: Our study provides evidence that TNF-α inhibition may weaken the ability of BMSCs in neurological functional recovery after SCI by activating the Wnt signaling pathway.